A comprehensive review of circRNA: from purification and identification to disease marker potential

Abstract

Circular RNA (circRNA) is an endogenous noncoding RNA with a covalently closed cyclic structure. Based on their components, circRNAs are divided into exonic circRNAs, intronic circRNAs, and exon-intron circRNAs. CircRNAs have well-conserved sequences and often have high stability due to their resistance to exonucleases. Depending on their sequence, circRNAs are involved in different biological functions, including microRNA sponge activity, modulation of alternative splicing or transcription, interaction with RNA-binding proteins, and rolling translation, and are a derivative of pseudogenes. CircRNAs are involved in the development of a variety of pathological conditions, such as cardiovascular diseases, diabetes, neurological diseases, and cancer. Emerging evidence has shown that circRNAs are likely to be new potential clinical diagnostic markers or treatments for many diseases. Here we describe circRNA research methods and biological functions, and discuss the potential relationship between circRNAs and disease progression.

Keywords: CircRNA; Database; Diseases; Marker.

Figure 1. Characteristics of different types of circRNA.

(A) mRNA: A class of single-stranded ribonucleic acids with genetic information transcribed from deoxyribonucleic acid (DNA). (B) Exon skipping event results in covalently splices and forms an ecRNA after the introns were removed. (C) The interaction between two RBPs can bridge two flanking introns together and form ecRNA, ElciRNA and mRNA. (D) RNA polymerase cleaves the intron from pre-mRNA to form an annulus, the circRNA formed in this manner is ciRNA.

Figure 2. Two different models of exon circularization of circRNA.

(A) Intron-pairing-driven circularization: during the formation of circRNA, an intron reverse complementary motif comprising GU-rich and C-rich elements is the key component to facilitate cyclization. (B) Lariat-driven circularization: the formation of circRNA is facilitated by the lariat structure. The complementary ALU flanking element which is repeated in the intron region competing for classical linear RNA splicing and the circularization is accelerated by reverse complementarity.

Figure 3. The five main functions of the circRNA.

(A) Regulating selective splicing or transcription: Stable circRNA and EIciRNAs are located in the nucleus, where they bind to RNA polymerase and promoting transcription; circRNA competes with pre-mRNA splicing to reduce the level of linear mRNA and excludes specificity from pre-mRNA by changing the composition of processed mRNA; (B) interaction with RBPs: circRNA binds with RBPs and ribonucleoprotein complexes and interfere with their functions. As a single circRNA can bind with multiple units of RBPs, they serve as stores of RBPs; (C) miRNA sponging activity: circRNA binds with miRNA and affecting the miRNA-dependent target gene suppression; (D) rolling circle translation: some circRNA can be translated into proteins by means of a roll loop amplification mechanism; (E) generation of pseudogenes: some circRNA are reverse transcribed into cDNA and integrated into the genome; however, the mechanism of integration is not yet clear.

Figure 4. The difference between linear RNA and circRNA primer design.

(A) FW is a forward primer with the b chain as template. The base sequence of synthesis is the original sequence of a. RV is a reverse primer with a chain as template, and the base sequence of synthesis is the original sequence of b. The sequence between FW and RV is high; (B) The original primers need to reverse: the synthetic primers are FW′ and RV′, where FW′ is the reverse complementary sequence of the RV primer, RV′ is the reverse complementary sequence of FW primer.